This invention relates to a novel process for obtaining pure 9-(2,6-dihalobenzyl)adenines from crude reaction mixtures containing substantial amounts of 3-(2,6-dihalobenzyl)adenines. Said crude reaction mixtures containing 9-(2,6-dihalobenzyl)adenines can be obtained by alkylating adenine by the process described in U.S. Ser. No. 766,326 filed Feb. 7, 1977 or U.S. Pat. No. 3,846,426. Said alkylated adenines are described in U.S. Pat. No. 3,846,426 as being useful in the treatment and prevention of coccidiosis.
Coccidiosis is a widespread poultry disease which is produced by infections of protozoa of the genus Eimeria which causes severe disorders in the intestines and ceca of poultry. Some of the most significant of these species are E. tenella, E. acervulina, E. necatrix, E. brunetti and E. maxima. This disease is generally spread by the birds picking up the infectious organism in droppings on contaminated litter or ground, or by way of contaminated food or drinking water. The disease is manifested by hemorrhage, accumulation of blood in the ceca, passage of blood in the droppings, weakness and digestive disturbances. The disease often terminates in the death of the animals, but the flow which survive severe infections have had their market value substantially reduced as a result of the infection. Coccidiosis is, therefore, a disease of great economic importance and extensive work has been done to find new and improved methods for controlling and treating coccidial infections in poultry.
9-(2,6-Dihalobenzyl)adenines, useful for the control and treatment of coccidial infections, are prepared by alkylation of adenine. Alkylation of adenine results primarily at the 9-position, but substantial amounts of the undesired 3-isomer and traces of the 7-isomer are also formed. The alkylation reaction and the structure of the main products i.e., the 3-isomer and 9-isomer are illustrated by the following equation: ##STR1## wherein X and X' are independently fluorine, chlorine or bromine.
After the reactants, adenine and .alpha.-chloro-2,6-dihalotoluene, are contacted with each other in the reaction medium, the product of the reaction, i.e., the 9-(dihalobenzyl)adenine, along with the other isomers that form, (e.g. the 3-isomer and to a lesser extent the 7-isomer) will either remain entirely in solution or precipitate on standing, depending upon the quantity of solvent utilized. Upon completion of the reaction, the reaction mixture may be cooled, e.g., to a temperature of about room temperature to precipitate a solid product or to precipitate a further quantity of this product. The product is then isolated in the usual manner, such as by filtration, and, if desired, purified by conventional methods such as washing with ethanol or water and recrystallizing from a suitable solvent such as acetic acid, aqueous acetic acid, dimethylformamide or dimethylsulfoxide. Swishing the product with dilute nitric acid or tetrafluoroboric acid (HBF.sub.4) also results in some purification.
The conventional purification methods recited above such as acetic acid recrystallization of crude 9-(2,6-dihalobenzyl)adenine produces a 99% pure product containing about 1% of the 3-isomer. Since it was learned that the 3-isomer gives a weak positive Ames test, an objective was set aimed at its virtual complete removal to levels less than 100 ppm. The process of the present invention is directed to methods for 3-isomer removal which involves a transalkylation reaction wherein the 2,6-dihalobenzyl group at the 3-position of adenine is removed as a carbenium ion by sulfuric acid and the carbenium ion allowed to react with a suitable carbenium ion trap. Alternatively, the crude 9-(2,6-dihalobenzyl)adenine may be subjected to a first step preliminary purification by extraction with dilute mineral acid before treatment with sulfuric acid as a second step. Furthermore, any conventional purification method recited above may be used as a preliminary first step to obtain partial purification before treatment with sulfuric acid. The sulfuric acid treatment process of the present invention produces 9-(2,6-dihalobenzyl)adenine containing &lt;100 ppm of 3-isomer in 92-94% yield over the two steps or in 96% yield in one step.